Formation of amyloid-like fibrils upon limited proteolysis of bovine α-lactalbumin

Bovine α-lactalbumin (α-LA) (10gL−1) was incubated with a protease from Bacillus licheniformis at pH 7.5 and 50°C. The reaction was biphasic consisting of an initial hydrolysis of intact α-LA and formation of dimers from large hydrolysis products within 60min followed by aggregation of dimers into aggregates of 500kDa. The aggregates consisted primarily of fibrillar strands with a diameter of 5nm. Formation of these strands was accompanied by a change in secondary structure towards higher β-sheet content and strong binding of thioflavin, features shared with amyloidal fibrils. The main components in these fibrils were fragments of 8.8 and 9.8kDa shown to occur in a monomer–dimer equilibrium. These fragments were identified and a molecular mechanism involving side-by-side assembly of dimers of these fragments into fibrils is proposed.